Inactivation of Escherichia coli by a combination of nisin,pulsed electric fields,and water activity reduction by sodium chloride

The effect of nisin combined with pulsed electric fields (PEF) and water activity reduction by sodium chloride (NaCl) on the inactivation of E. coli in simulated milk ultrafiltrate media was studied with a Doehlert design and a response surface method. The reduction of water activity from 0.99 to 0.95 by the addition of NaCl (without any other hurdle) did not affect E. coli viability of approximately 10(8) CFU/ml. A reduction in PEF effectiveness occurred when the NaCl concentration was increased because of an increase in conductance, which reduced the pulse decay time. In cells submitted to PEF nisin activity was decreased, probably as a consequence of the nonspecific binding of nisin to cellular debris or the emergence of new binding sites in or from cells. However, the lethal effect due to nisin was reestablished and further improved when water activity was reduced to 0.95. A synergistic effect was evidenced when low-intensity PEF were applied. Decreasing water activity to 0.95 and applying PEF at 5 kV/cm (a nonlethal intensity when no other hurdle is used) with the further addition of nisin (1,200 IU/ml) resulted in a 5-log cycle reduction of the bacterial population.     

Effect of chelators and nisin produced in situ on inhibition and inactivation of gram negatives

The ability of chelators and nisin generated in situ to inhibit and inactivate E. coli and other gram negatives in a model substrate was investigated. The effect of various chelators and different concentrations of exogenous nisin on inhibition of E. coli in broth medium showed that only EDTA and pyrophosphates were able to cause appreciable inhibition of E. coli by nisin. In a broth where L. lactis NCFB 497 produced nisin in a concentration of 250-300 IU/ml, pyrophosphates were unable to inactivate E. coli. Under the same conditions, addition of EDTA led to inactivation of E. coli at neutral and slightly acidic pH only. A cocktail of strains of E. coli was less sensitive than E. coli ATCC 25922 alone. Pseudomonas aeruginosa was more sensitive and salmonellae more resistant. EDTA also caused a slight reduction in the L. lactis population and its biochemical activity as regards pH drop and acid production. Some of the inhibition of E. coli could be ascribed to the physical presence of Lactococcus cells rather than their metabolites excreted into the medium. Failure to observe any inhibition in fermented broths at their natural pH (4.0) was ascribed to the poor chelating power of EDTA under acid conditions.     

Inhibitory combinations of nisin, sodium chloride, and pH on Listeria monocytogenes ATCC 15313 in broth by an experimental design approach

The influence of pH (5.0-8.2), NaCl concentrations (0-6% w/v), and incubation time (0-24 h) on the inhibitory activity of nisin (0-100 I.U./ml) against Listeria monocytogenes (10(3) cfu/ml) was studied using the Doehlert experimental design and was confirmed by kinetic experiments. Predicted values were in agreement with experimental values. Experiments were carried out at 22 degrees C in reconstituted TSB-YE1 broth with or without NaCl. Nisin had an immediate pH-dependent bactericidal effect, which increased with decreasing pH values. In modified TSB-YE1 broth without NaCl, the bactericidal efficacy of nisin (50 I.U./ml) was maximum at pH 6.6, with no L. monocytogenes survivors until 120 h at 22 degrees C. Nisin (50 I.U./ml) action decreased in the presence of NaCl, with a minimal inhibitory effect between 2 and 4%. This partially protective effect was cancelled at higher levels of nisin.     

炝蟹大肠菌群的控制

研究了炝蟹中大肠菌群的受各种因子(aw、T、Pres、pH、Eh、腌制时间)的影响情况。实验结果表明,渗透压的变化对大肠菌群的生长影响很大,当盐浓度为10%开始受到抑制并随着培养时间的延长效果增大;pH值为7时大肠杆菌的生长良好,强酸较强碱不利于其生长;保鲜剂具有较好的抑制效果;真空影响不大。使炝蟹盐度为3%和采用保鲜剂控制微生物的生长,能达到国家指定的腌制品的标准。     

Resistance of Escherichia coli and Salmonella against nisin and curvacin A.

We have determined the effects of the following factors on the resistance of Gram-negative bacteria against nisin and curvacin A: (i) chemotype of the lipopolysaccharide (LPS), (ii) addition of agents permeabilizing the outer membrane, (iii) the fatty acid supply of the growth medium, and (iv) the adaptation to acid and salt stress. Bacteriocin activity was determined against growing and resting cells as well as protoplasts. All smooth strains of Escherichia coli and Salmonella enterica serovar Typhimurium were highly resistant towards the bacteriocins, whereas mutants that possess the core of the LPS, but not the O antigen, as well as deep rough LPS mutants were sensitive. Antibiotics with outer membrane permeabilizing activity, polymyxin B and polymyxin B nonapeptide, increased the sensitivity of smooth E. coli towards nisin, but not that of deep rough mutants. Incorporation of 1 g l(-1) of either oleic acid or linoleic acid to the growth media greatly increased the susceptibility of E. coli LTH1600 and LTH4346 towards bacteriocins. Both strains of E. coli were sensitive to nisin and curvacin A at a pH of less than 5.5 and more than 3% (w/v) NaCl. Adaptation to sublethal pH or higher NaCl concentrations (pH 4.54 and 5.35 or 4.5% (w/v) NaCl) provided only limited protection against the bacteriocidal activity of nisin and curvacin A. Adaptation to 4.5% (w/v) NaCl did not result in cross protection to bacteriocin activity at pH 4.4, but rendered the cells more sensitive towards bacteriocins.     

Survival of Escherichia coli O157:H7 in dry fermented sausages containing micro-encapsulated probiotic lactic acid bacteria

Escherichia coli O157:H7 is capable of surviving the rigorous processing steps during the manufacture of dry fermented sausages. The effect of adding two probiotic organisms, Lactobacillus reuteri and Bifidobacterium longum as co-cultures with the meat starter cultures Pediococcus pentosaceus and Staphylococcus carnosus on the viability of E. coli O157:H7 in dry fermented sausages was studied. A 5 strain cocktail of E. coli O157:H7 was added at 7.4 log cfu/g to the sausage batter and challenged with either or both Lb. reuteri or B. longum before or after they were micro-encapsulated. Sausages were fermented at < or = 26 degrees C and 88% relative humidity (RH) followed by drying at 75% RH and 13 degrees C for 25 d. The pH, water activity (aw), protein, moisture, and numbers of all inoculated organisms were monitored during processing. The pH and aw decreased from 5.7 and 0.98 to 4.9 and 0.88 at the end of fermentation and drying, respectively. These processes reduced E. coli O157:H7 by 1.0 and 0.7 log cfu/g at the end of fermentation and drying, respectively. Unencapsulated Lb. reuteri with or without B. longum reduced E. coli O157:H7 by 3.0 log cfu/g and B. longum caused a 1.9 log cfu/g reduction. While micro-encapsulation increased survival of Lb. reuteri and B. longum, it reduced their inhibitory action against E. coli O157:H7.     

Combined effect of nisin and pulsed electric fields on the inactivation of Escherichia coli

The Doehlert design was applied in order to investigate the combined effect of nisin and high voltage pulsed electric fields (PEF) on the inactivation of Escherichia coli in simulated milk ultrafiltrate media. Nisin alone was totally inactivated by PEF, but in the presence of bacterial cells a protective effect was observed. However, the effectiveness of nisin was still decreased when bacterial cells were subjected to the combined treatment. In spite of this phenomenon, an almost additive response emerged as a consequence of the combined treatment. A 4-log cycle reduction may be accomplished with around 1,000 IU/ml (7.15 microM) of nisin and three pulses of 11.25 kV/cm or 500 IU/ml for five pulses of the same intensity. The observed efficacy arising from the combination of both treatments suggests the possibility of using PEF for improving the action spectrum of natural antimicrobials.     

辅酶Q0的抑菌作用及稳定性研究

研究辅酶Q0对6种常见食源性致病菌的抑制效果以及辅酶Q0在温度、pH、可见光和超声下的抑菌稳定性,并采用平板涂布法进一步探究其在食品体系中的抑菌效果。结果表明:辅酶Q0具有良好的抑菌活性,其对金黄色葡萄球菌、单增李斯特菌、副溶血性弧菌的最小抑菌浓度(minimum inhibitory concentration,MIC)分别为0.031、0.025、0.009 mg/m L,对大肠杆菌、鼠伤寒沙门氏菌和阪崎肠杆菌的MIC均为0.250 mg/m L。温度、超声和可见光对辅酶Q0的抑菌活性无明显影响;介质的pH会极大地影响辅酶Q0的抑菌活性。食品体系抑菌实验表明,辅酶Q0在巴氏灭菌奶中对金黄色葡萄球菌的抑制杀灭作用优于在TSB中对金葡菌的抑杀作用;辅酶Q0对豆浆中腐败菌的抑制效果优于乳酸链球菌素。该研究为辅酶Q0在食品、医药行业的利用提供了一定的理论依据。      

常用防腐剂对牛乳铁蛋白抗菌活性的影响

乳铁蛋白的抗菌活性常受到其他共存成分的影响。本文研究了常用防腐剂对乳铁蛋白抗菌活性的影响。在研究过程中,以大肠杆菌为实验菌株,进行了实验研究。研究表明,在牛肉膏蛋白胨培养基中,pH6.0,菌浓为106CFU/ml的条件下,乳铁蛋白对大肠杆菌的最小抑菌质量浓度为3mg/ml;苯甲酸钠、山梨酸钾、乳酸链球菌素、对羟基苯甲酸等分别与乳铁蛋白共同作用时,对乳铁蛋白的抗菌活性都有明显的影响。其中,山梨酸钾和苯甲酸钠减弱了乳铁蛋白的抗菌活性,乳酸链球菌素和对羟基苯甲酸丁酯加强了乳铁蛋白的抗菌活性。     

Specific Bifidobacterium strains isolated from elderly subjects inhibit growth of Staphylococcus aureus

Cell-free, pH-controlled supernatants of thirty-eight Bifidobacterium strains isolated from healthy elderly subjects were subjected to antimicrobial activity assay. Bioluminescent indicator strains Staphylococcus aureus RN4220, Escherichia coli K-12, and Salmonella enterica serovar Typhimurium ATCC 14028 were used as targets of antimicrobial activity. The effect of nutrient depletion on the inhibition was eliminated with spent-culture controls. Three out of thirty-eight Bifidobacterium strains were capable of inhibiting the growth of S. aureus. The inhibition was equal to 23.2+/-19.1% to 50.4+/-26.7% of the inhibition caused by 50 IU/ml nisin. Reuterin-producing positive strain Lactobacillus reuteri SD2112 was capable of 86.0+/-24.6% inhibition, but Bifidobacterium lactis Bb-12, a known probiotic strain, showed no inhibition. None of the strains was capable of inhibiting the growth of E. coli or S. enterica. The observed inhibition by bifidobacteria was related to hydrogen peroxide formation and possible production of heat-stable proteinaceous compounds. The results suggest that production of antimicrobial substances other than organic acids is not common among Bifidobacterium strains typical of elderly subjects. However, specific strains were identified which showed considerable inhibitory activity against S. aureus.     

国产Nisin部分特性的研究

研究温度、pH、蛋白酶、保存条件对国产Nisin抑菌活性的影响,以及抑菌谱等,结果表明温度、pH会显著影响Nisin活性,在酸性条件下,Nisin对温度较稳定,随PH值的增加,温度越高,Nisin活性下降越显著,温度越低越有利于保存,干制品比溶液易保存,国产Nisin对胃蛋白酶不敏感,但对胰蛋白酶敏感,国产Nisin对革兰氏阳性细菌有抑制作用,而对革兰氏阴性细菌、霉菌和酵母没有作用。     

肉源乳酸菌抑菌特性分析

为了筛选有抑菌活性的乳酸菌并分析其抑菌的物质基础,利用牛津杯琼脂扩散法,分析了19株从牧区风干肉制品中分离得到的肉源乳酸菌发酵上清液对大肠杆菌和金黄色葡萄球菌的抑菌能力,同时从中分别选取对大肠杆菌和金黄色葡萄球菌抑制能力有显著差异的乳酸菌各3株,对其上清液中的抑菌物质基础和抑菌能力影响因素进行了分析。结果表明19株乳酸菌中菌株F19对大肠杆菌的抑菌活性最强,抑菌圈直径为(15.07±0.55) mm,而F11发酵上清液对大肠杆菌完全没有抑制效果,菌株F16对金黄色葡萄球菌的抑菌活性最强,抑菌圈直径为(14.47±0.38) mm,而F6、F2发酵上清液对金黄色葡萄球菌完全没有抑制效果;抑菌的物质基础分析表明,酸性环境在抑菌过程中占主导地位,细菌素与过氧化氢发挥协同作用;对其发酵上清液进行不同pH以及不同温度处理后发现,其抑菌活性在pH<5.0时随pH的降低而显著增强(P<0.05),当pH分别升高至5.0、6.0、7.0时试验乳酸菌发酵上清液均不具有抑菌能力,F18发酵上清液分别在60、80、100、121 ℃热处理一定时间后其抑菌能力与对照组相比仍然无明显差异(P>0.05),F11发酵上清液在100 ℃热处理30 min后抑菌能力显著减弱(P<0.05),其余实验菌株发酵上清液温度达到60 ℃以上,抑菌能力随热处理温度提高而显著(P<0.05)下降。本研究发现了乳酸菌发酵上清液的抑菌效果具有特异性,抑菌物质主要是酸性物质,其次是细菌素和过氧化氢;抑菌能力受pH和温度影响,同时具有特异性。研究结果可为优质菌种资源开发提供理论指导与技术支持。     

Antimicrobial effect of natural preservatives in a cooked and acidified chicken meat model

The inhibitory effect of Microgard 100, Microgard 300, nisin, Alta 2002, Perlac 1902, sodium lactate and essential oil of mustard on microorganisms experimentally inoculated was screened in an acidified chicken meat model (pH = 5.0) and stored for 2 weeks at a none restrictive growth temperature of 22 degrees C. All antimicrobials tested were used at the highest concentration recommended by their manufacturer. Sausage batter made with mechanically deboned chicken was inoculated with a mixed culture of Escherichia coli ATCC 25922, Brochothrix thermosphacta CRDAV452, and a protective culture Lactobacillus alimentarius BJ33 (FloraCan L-2). A final cell concentration of 3-4 log CFU g (-1) was targeted after cooking at a core temperature of 55 degrees C for each microorganism in order to assess cell count variation effectively. Composition, water activity (a(w)), pH and redox potential of the sausage model was also evaluated. The E. coli population decreased steadily during storage and was close or below detection level (< 1 log CFU g (-1)) for all treatments, including the control, after 14 days. Sodium lactate was most effective against B. thermosphacta; population was 4 log lower than the control after 14 days of storage. When essential oil of mustard was used, aerobic mesophilic bacteria and lactic acid bacteria were significantly lower than the control after 2 days of storage (P < or = 0.05). The other antimicrobial agents tested had no significant effect on the aerobic mesophilic bacteria, E. coli, B. thermosphacta and lactic acid bacteria counts, when compared to the control.     

产细菌素屎肠球菌E6的特性分析及发酵条件优化

研究了屎肠球菌E6抑菌活性的生物学特性,并通过响应面法优化其发酵条件。结果表明,屎肠球菌E6产蛋白质类细菌素,能够抑制单增李斯特氏菌、大肠杆菌、金黄色葡萄球菌和枯草芽孢杆菌生长,在pH3.0～7.0条件下有明显抑菌活性,60～121℃热处理20min后仍具有抑菌活性。通过Box-Behnken实验设计优化屎肠球菌E6发酵条件为培养时间36.0h,培养温度31.0℃,培养基pH5.1。在此条件下,发酵上清液抑菌圈直径可达20.17mm,较优化前提高了27.2%。      

Physical and Chemical Properties of Edible Films Containing Nisin and Their Action Against Listeria Monocytogenes

ABSTRACT: The effects of hydrophobicity/hydrophilicity of edible films against Listeria monocytogenes strain V7 by various nisin concentrations (4.0 - 160 IU/film disk) and pH values ranging from 2.0 to 8.0 were determined and the mechanical properties and water vapor permeability of films prepared with or without nisin were compared. Surface hydrophobicities (446, 282, 232 and 142, respectively) of whey protein isolates, soy protein isolates, egg albumen and wheat gluten were determined. As the nisin concentration increased, the amount of inhibition progressively increased in all tested films. Using nisin, edible films with higher hydrophobicity values of 280 to 450 units under an acidic environment exerted a greater inhibitory effect against L. monocytogenes. Different interactions of nisin with the proteins of the different films resulted in variation in the mechanical properties and water vapor permeabilities of the films tested.     

Inactivation of Salmonella typhimurium and Escherichia coli O157:H7 in apple juice by a combination of nisin and cinnamon

Pasteurized apple juice with nisin (0, 25, 50, 100, and 200 ppm, wt/vol) and cinnamon (0 and 0.3%, wt/vol) was inoculated with Salmonella Typhimurium and Escherichia coli O157:H7 at 10(4) CFU/ml and stored at 5 and 20 degrees C. Counts on tryptic soy agar (TSA), selective medium (xylose Lysine desoxycholate agar for Salmonella Typhimurium, and MacConkey sorbitol agar for E. coli O157:H7), and thin agar layer (TAL) were determined at 1 h and 1, 3, 7, and 14 days. The TAL method (selective medium overlaid with TSA) was used for recovery of sublethally injured cells. The pathogens were gradually inactivated by the acidic pH of apple juice. Nisin and cinnamon greatly contributed to the inactivation. The killing effect was more marked at 20 degrees C, with counts in all treated samples being undetectable by direct plating in 3 days for Salmonella Typhimurium and 7 days for E. coli O157:H7. Thus, several factors influenced the decrease in counts: low pH, addition of nisin and cinnamon, and storage temperature. The TAL method was as effective as TSA in recovering injured cells of the pathogens. The combination of nisin and cinnamon accelerates death of Salmonella Typhimurium and E. coli O157:H7 in apple juice and so enhances the safety of the product.     

Stability and antimicrobial efficiency of eugenol encapsulated in surfactant micelles as affected by temperature and pH

Growth inhibition of four strains of Escherichia coli O157:H7 (H1730, F4546, 932, and E0019) and Listeria monocytogenes (Scott A, 101, 108, and 310) by eugenol encapsulated in water soluble micellar nonionic surfactant solutions (Surfynol 485W) adjusted to pH 5, 6, and 7 and incubated at 10, 22, and 32 degrees C was determined. Concentrations of eugenol ranged from 0.2 to 0.9% at a surfactant concentration of 5%. Antimicrobial activity was assessed using a microbroth dilution assay. Eugenol encapsulated in surfactant micelles inhibited both microorganisms at pH 5, 6, and 7. At pH 5, some inhibition occurred in the absence of eugenol, i.e., by the surfactant itself (optical density at 24 h for L. monocytogenes = 0.07 and optical density at 24 h for E. coli O157:H7 = 0.09), but addition of >0.2% eugenol led to complete inhibition of both microorganisms. Inhibition of L. monocytogenes and E. coli O157:H7 decreased with increasing pH, that is, the minimum inhibitory concentration was 0.2, 0.5, and 0.5% of micellar encapsulated eugenol solutions at pH 5, 6, and 7, respectively. The encapsulated essential oil component in surfactant micelles was effective at all three temperatures tested (10, 22, and 32 degrees C), indicating that the activity of encapsulated eugenol was not affected by high or low (refrigeration) temperatures. Overall, strains of E. coli O157:H7 were more sensitive than strains of L. monocytogenes. Improved activity was attributed to increased solubility of eugenol in the aqueous phase due to the presence of surfactants and improved interactions of antimicrobials with microorganisms.     

不同发酵剂和工艺环节对发酵牛肉调味基料抑菌性的影响

为研究不同发酵剂和工艺条件对发酵牛肉调味基料（fermented beef flavorings,FBF）抑菌性的影响,以金黄色葡萄球菌（Staphylococcus aureus）、鼠伤寒沙门氏菌（Salmonella typhimurium）、铜绿假单胞菌（Pseudomonas aeruginosa）、大肠杆菌（Escherichia coli）和乙型副伤寒沙门氏菌（Salmonella paratyphi B）为目标菌,添加10% FBF于目标菌中,37 ℃培养12 h,计算其对不同目标菌的抑制率。设计3 组实验：CK组（热压浸提-酶解-美拉德反应）;2）反应Ⅰ组（热压浸提-酶解-发酵）,在发酵环节接种15 种商业复合菌或单株菌;3）反应Ⅱ组（热压浸提-酶解-发酵-美拉德反应）,在反应Ⅰ的基础上继续进行美拉德反应。结果表明：不同工艺制得的处理组FBF,其抑菌能力从强到弱依次为反应Ⅱ组＞反应Ⅰ组＞CK组;CK组对P. Aeruginoosa和S. paratyphi B无抑菌能力,对E. coli的抑制率最高（61%）,其次是S. aureus（20%）和S. typhimurium（11%）;反应Ⅰ制得的FBF除对P. Aeruginoosa无抑制能力外,对其余4 株目标菌的抑制率均提高,其中,以清酒乳杆菌为发酵剂制得的FBF对S. typhimurium的抑制率可达55%;反应Ⅱ制得的15 种FBF对5 株目标菌均有较强的抑菌性,以清酒乳杆菌为发酵剂制得的FBF对S. typhimurium的抑制率达85%,对E. coli的抑制率达53%,以WBL-45为发酵剂制得的FBF对E. coli的抑制率达100%,对S. paratyphi B的抑制率达85%。最终选定WBL-45、清酒乳杆菌为制作FBF的发酵剂,采用热压浸提-酶解-发酵-美拉德反应工艺制作FBF,得到的产品抑菌性较强。     

Enhanced inhibition of Escherichia coli O157:H7 by lysozyme and chelators

This study examined the effects of three chelating agents (EDTA, disodium pyrophosphate [DSPP], and pentasodium tripolyphosphate [PSTPP]) on the inhibition of the growth of Escherichia coli O157:H7 by lysozyme. The objective of this study was to identify replacement chelators that exhibit synergistic properties similar to those of EDTA. The inhibitory effects of EDTA at 300 to 1,500 microg/ml and of DSPP and PSTPP at 3,000 to 15,000 microg/ml in combination with lysozyme at 200 to 600 microg/ml for up to 48 h at pHs of 6.0, 7.0, and 8.0 on four strains of E. coli O157:H7 was studied with the use of a microbroth dilution assay. The addition of EDTA enhanced lysozyme's inhibitory effect on strains of E. coli O157:H7. EDTA at > or = 300 microg/ml combined with lysozyme at 200 to 600 microg/ml was sufficient to inhibit the growth of the strains at pHs of 6.0 and 8.0. At pH 7.0, lysozyme at 200 to 600 microg/ml and EDTA concentrations of > or = 1,000 microg/ml were effective in inhibiting three of the four strains. DSPP at pH 6.0 was inhibitory at > or = 10,000 microg/ml when combined with lysozyme at 200 to 300 microg/ml. In contrast, PSTPP increased the inhibitory activity of lysozyme more effectively at pH 8.0. Lysozyme at 200 to 600 microg/ml was effective against two strains of E. coli O157:H7 when used in conjunction with PSTPP at > or = 5,000 microg/ml. The remaining strains were inhibited by PSTPP at > or = 10,000 microg/ml. Our results indicate that inhibition occurred with each lysozyme-chelator combination, but the concentrations of phosphates required to increase the antimicrobial spectrum of lysozyme against E. coli O157:H7 were higher than the EDTA concentrations required to achieve the same effect.     

Enhancement of nisin, lysozyme, and monolaurin antimicrobial activities by ethylenediaminetetraacetic acid and lactoferrin

A microtiter plate assay was employed to systematically assess the interaction between ethylenediaminetetraacetic acid (EDTA) or lactoferrin and nisin, lysozyme, or monolaurin against strains of Listeria monocytogenes, Escherichia coli, Salmonella enteritidis, and Pseudomonas fluorescens. Low levels of EDTA acted synergistically with nisin and lysozyme against L. monocytogenes but EDTA and monolaurin interacted additively against this microorganism. EDTA synergistically enhanced the activity of nisin, monolaurin, and lysozyme in tryptic soy broth (TSB) against two enterohemorrhagic E. coli strains. In addition, various combinations of nisin, lysozyme, and monolaurin with EDTA were bactericidal to some gram-negative bacteria whereas none of the antimicrobials alone were bactericidal. Lactoferrin alone (2000 microg ml(-1)) did not inhibit any of the bacterial strains, but did enhance nisin activity against both L. monocytogenes strains. Lactoferrin in combination with monolaurin inhibited growth of E. coli O157:H7 but not E. coli O104:H21. While lactoferrin combined with nisin or monolaurin did not completely inhibit growth of the gram-negative bacteria, there was some growth inhibition. All combinations of EDTA or lactoferrin with antimicrobials were less effective in 2% fat UHT milk than in TSB. S. enteritidis and P. fluorescens strains were consistently more resistant to antimicrobial combinations. Resistance may be due to differences in the outer membrane and/or LPS structure.